Protected α-amino-α-ureido(thioureido)phenylacetic acids

ABSTRACT

Cephalosporins with a α-amino-(ureidophenyl)acetamido substituent at position 7 are prepared by acylation of a 7-aminocephalosporin with a derivative of α-amino-p(or m)-ureidophenylacetic acid. These compounds are antibacterial agents.

This is a division of application Ser. No. 357,763 filed May 7, 1973,now U.S. Pat. No. 4,007,173.

This invention relates to new cephalosporin compounds with improvedproperties. In particular, the new compounds have anheterocyclicthiomethyl group at position 3 and a ureidophenylglycylsubstituent at position 7 of the cephem nucleus. These compounds haveantibacterial activity when administered either orally or parenterally.

Since the discovery of the cephalosporins as a new group ofantibacterials, a wide variety of semi-synthetic analogs has beenprepared. Although many derivatives have been found that display a highdegree of activity when administered parenterally, the discovery ofcephalosporins that have oral activity has presented a difficultproblem. Cephaloglycine, U.S. Pat. No. 3,560,489; cephalexin, U.S. Pat.No. 3,507,861; and cephradine, U.S. Pat. No. 3,485,819 are the onlycompounds that have commercial use in this area. We have now discovereda new series of cephalosporins that have oral activity.

The compounds of this invention are defined by the following formula:##STR1## wherein: THE R₂ NCXNR group is at the para or meta position;

R is hydrogen or lower alkyl of 1-4 carbon atoms;

X is oxygen or sulfur;

A is hydrogen, acetoxy, methylthio, methoxy, azido, or SHet; and

Het is a 5 or 6-membered heterocyclic ring containing carbon and 1-4atoms selected from the group consisting of N, O, and S, unsubstitutedor substituted with one or two substituents selected from the groupconsisting of C₁ -C₆ alkyl, C₁ -C₆ alkoxy, allyloxy, oxide, carboxamido,carboxyl, carbalkoxy of C₁ -C₆, halogen, mercapto, methylthio,trifluoromethyl, hydroxy, amino, alkylamino and dialkylamino, eachundefined alkyl having 1-6 carbon atoms.

Het includes N-oxide derivatives of the heterocyclic systems named wheresuch derivative is possible, for example, pyridyl-N-oxide.

Preferred compounds are those where Het is tetrazolyl, triazolyl,thiadiazolyl, oxadiazolyl, diazolyl, pyridyl, pyrimidyl and pyrazinyl.

Also preferred are compounds where R is hydrogen. Particularly preferredcompounds are those where Het and R are as preferred above and X isoxygen.

Due to the presence of the amino and carboxylic acid groups, thecompounds of this invention can exist as the zwitterion or an acid orbase salt. These salts are prepared by standard methods using a widevariety of nontoxic pharmaceutically acceptable acids and bases that areknown in the art.

The asymmetric carbon in the ureidophenylglycine side chain gives riseto optical isomers, of which the diastereomers having theD-configuration in the sidechain are preferred; however, those havingthe L-configuration and the diastereomeric mixtures derived fromacylations using racemic side chain acids are within the scope of theinvention.

The compounds of this invention are prepared by acylation of theappropriate 7-aminocephalosporin nucleus with the appropriateureidophenylglycine. The carboxylic acid group is activated by any ofthe standard methods such as mixed anhydride, acid chloride, oractivated ester. In addition, a coupling reagent, for exampledicyclohexylcarbodiimide, can be used provided that the carboxyl groupon the cephem nucleus is protected with an easily removable protectinggroup. The amino group of the glycine moiety must also be protectedduring the acylation. Examples of protecting groups known in the art aret-butoxycarbonyl, trichloroethoxycarbonyl, benzyloxycarbonyl, the methylacetoacetate adduct, or similar groups commonly used in peptidesynthesis.

Alternatively, the compounds where A is SHet may be prepared byacylating 7-aminocephalosporanic acid with the N-protectedureidophenylglycine and then displacing the acetoxy group with thedesired heterocyclicthiol. Removal of the protecting group gives theproduct compounds of this invention.

The starting materials are known, prepared by known methods, ordescribed herein. Ureidophenylglycines, both the meta and paracompounds, are prepared from p-aminophenylglycine andm-aminophenylglycine (U.S. Pat. No. 3,479,339) by first protecting theglycine amino group and then reacting the phenyl amino group withcyanate ion. The thioureidophenylglycines are similarly prepared byusing t-butyl isothiocyanate and hydrolyzing the t-butyl group. Themonoalkylureido compounds are prepared by substituting an alkylisocyanate for the cyanate ion or by reacting N-alkylaminophenylglycinewith cyanate ion after first protecting the glycine amino group. Thedialkylureido derivatives are prepared by substituting dialkylcarbamylchloride for the cyanate ion. The7-amino-3-heterocyclicthiomethyl-3-cephem-4-carboxylic acids areprepared by displacing the acetoxy group of 7-aminocephalosporanic acid(7-ACA) with a heterocyclicthiol by the method disclosed in U.S. Pat.No. 3,516,997.

The compounds have antibacterial activity against both Gram-positive andGram-negative organisms and are therefore useful for the treatment orprevention of bacterial infections. These compounds are particularlyuseful since the antibacterial activity in animals is observed when theyare administered either parenterally or orally. Minimum inhibitoryconcentrations (MIC) were determined using the standard tube dilutionmethod.3-Methyl-7-(α-amino-p-ureidophenylacetamido)-3-cephem-4-carboxylic acid(93916) had MIC's ranging from 3.1 to greater than 200 μg/ml when testedagainst a variety of bacteria. In addition,3-(5-methyl-1,3,4-thiadiazol-2-ylthiomethyl)-7-(α-amino-p-ureidophenylacetamido)-3-cephem-4-carboxylicacid (88062) had MIC's ranging from 0.4 to greater than 200 μg/ml. Thein vivo activity of these compounds was demonstrated by administeringthe compounds to mice infected with E. coli and Kleb. pneumonia. Theresults from both oral and subcutaneous administration are reported inTable 1 as ED₅₀ in mg/kg along with those of a standard, cephalexin.

                  TABLE 1                                                         ______________________________________                                                 E. coli      K. pneumo.                                              Compound*  sc       po        sc     po                                       ______________________________________                                        93916      37       45        22     19                                       35236      7.5      25        11     45                                       88026      2        4         3      2                                        66036      <3       5         5      5                                        56136      4        7         5      7                                        66136      <3,2     9,6       <3,2   10,3                                     Cephalexin 15       25        25     16                                       ______________________________________                                         *See Table 2 for structures                                              

                  TABLE 2                                                         ______________________________________                                         ##STR2##                                                                     Compound     R            A                                                   ______________________________________                                        93916        p-ureido     hydrogen                                            35236        p-ureido     acetoxy                                             88026        p-ureido     5-methyl-1,3,4-                                                               thiadiazol-2-                                                                 ylthio                                              66036        p-ureido     1-methyltetrazol-                                                             5-ylthio                                            56136        m-ureido     5-methyl-1,3,4-                                                               thiadiazol-2-                                                                 ylthio                                              66136        m-ureido     1-methyltetrazol-                                                             5-ylthio                                            ______________________________________                                    

The compounds are formulated in the same manner as other cephalosporins.They are administered parenterally as sterile aqueous solutions ororally as tablets, capsules, or suspensions. The amount given varieswith age, size, and condition of the subject as well as the severity ofthe infection.

The following examples are presented to illustrate the invention but arenot to be viewed as limiting the scope thereof.

EXAMPLE 1 α-t-Butoxycarbonylamino-p-ureidophenylacetic acid

A mixture of p-aminophenylglycine (27.73 g, 0.17 mol),t-butyl-2,4,5-trichlorophenylcarbonate (46.94 g, 0.23 mol),triethylamine (67.25 ml), t-butyl alcohol (165 ml), and water (110 ml)were stirred and heated at 62° for 2 hours. The alcohol was removed invacuo at 40° and the residue was diluted with water (190 ml), acidifiedto pH 3 with 40% H₃ PO₄ and then extracted with ethyl acetate. Thecombined extracts were washed with 5% NaHCO₃ (3 × 175 ml). The aqueouswashings were cooled, acidified to pH 3.0 with 40% H₃ PO₄, and extractedwith ethyl acetate. The dried extracts were evaporated to give theN-t-butoxycarbonyl derivative. Additional product was obtained from anemulsion which formed during the extraction. Total yield was 14.5 g.

The above product (8.18 g, 0.03 mol) was reacted with potassium cyanate(3.3 g, 0.04 mol) in acetic acid (8 ml) and water (105 ml) for 8.5 hoursat 55°. The reaction solution was cooled, acidified to pH 1 with 3 N HCland extracted with ethyl acetate. The dried extracts were evaporated invacuo to give the title compound: mp 137°-139°(dec).

EXAMPLE 23-(5-Methyl-1,3,4-thiadiazol-2-ylthiomethyl)-7-(α-amino-p-ureidophenylacetamido)-3-cephem-4-carboxylicacid

The product from Example 1 (3.1 g, 0.01 mol) was dissolved in drytetrahydrofuran (60 ml) and then triethylamine (1.4 ml) andN-methylmorpholine (2 drops) were added. The resulting solution wascooled to -15° and i-butyl chloroformate (1.3 ml, 0.01 mol) was added.After stirring the reaction for 25 minutes at -10°, a cold solution of7-amino-3-(5-methyl-1,3,4-thiadiazol-2-ylthiomethyl)-3-cephem-4-carboxylicacid (3.44 g, 0.01 mol) in a mixture of 50% aqueous tetrahydrofuran (80ml) and triethylamine (1.4 ml) was added. Additional triethylamine wasadded to adjust the reaction solution to pH 6.8-7. The reaction wasstirred at room temperature for 3 hours followed by evaporation of theTHF. The aqueous solution was extracted with ethyl acetate, which wasdiscarded, and then adjusted to pH 1 with 3N HCl while stirring withethyl acetate. Phases were separated and the aqueous phase wasreextracted with ethyl acetate. The combined extracts were dried andevaporated to the solid N-protected derivative. This product waspurified by dissolving in methanol (50 ml) and then diluting with ether(500 ml).

The N-protected derivative (1.12 g) was added to ice coldtrifluoroacetic acid (12 ml) and was stirred without further cooling for7 minutes. The solvent was removed in vacuo and ether was added to theresidue. The solid salt was collected, stirred with water, and filteredfrom the insoluble material. The aqueous solution was treated with basicion exchange resin (Amberlite IR-45) until pH 4.7 was obtained and thenfiltered. The title product was obtained by lyophization of the watersolution: mp < 300°.

EXAMPLE 33-Methyl-7-(α-amino-p-ureidophenylacetamido)-3-cephem-4-carboxylic acid

To a solution of the product from Example 1 (4.8 g, 0.0154 mol) andt-butyl 7-amino-3-desacetoxycephalorporanate (4.12 g, 0.016 mol) in drytetrahydrofuran (100 ml) was added dicyclohexylcarbodiimide (3.5 g,0.017 mol). The reaction is stirred at room temperature for 2 hours andthen the solid urea filtered off. The filtrate was evaporated to a foamwhich was triturated with ether: ethyl acetate to give 4.5 g of thet-butyl ester of the title product.

The ester (4.4 g) was stirred with ice cold trifluoroacetic acid (45 ml)with further cooling in ice for 1 hour. The reaction was evaporated,ether was added to the residue, and the salt was collected. The salt wasdissolved in water and treated with basic ion-exchange resin (AmberliteIR-45) until pH 4.75 was obtained. After filtration, the aqueoussolution was lyophilized to give the desired product: mp > 300°.

EXAMPLE 43-(1-Methyltetrazol-5-ylthiomethyl)-7-(α-amino-p-ureido-phenylacetamido)-3-cephem-4-carboxylicacid

When 7-amino-3-(1-methyltetrazol-5-ylthiomethyl)-3-cephem-4-carboxylicacid (3.28 g, 0.01 mol) was acylated with the product of Example 1 (3.1g, 0.01 mol) according to the procedure of Example 2, the title compoundwas obtained: mp > 300°.

EXAMPLE 53-(1,2,3-Triazol-4-ylthiomethyl)-7-(α-amino-p-ureidophenylacetamido)-3-cephem-4-carboxylicacid

To a stirred solution of α-t-butoxycarbonylamino-p-ureidophenylaceticacid (10.1 g, 0.0375 mol) in dry THF (150 ml) is added triethylamine(5.2 ml, 0.0375 mol). The mixture is cooled to -10° and then isobutylchloroformate (4.92 ml, 0.0375 mol) is added dropwise over a 10 minuteperiod. The reaction mixture is stirred at -10° for 70 minutes and thena cold solution of 7-ACA (10.1 g, 0.0375 mol) in 50% aqueous THF (140ml) and triethylamine (6.75 ml, 0.0487 mol) is added over a 15 minuteperiod. The reaction is stirred at -5° to 0° for 1 hour and at roomtemperature for 2 hours. The organic solvents are evaporated and water(150 ml) is added to the aqueous residue. The solution is extracted withethyl acetate and the aqueous phase is separated, covered with freshethyl acetate, acidified to pH 2.8, and filtered. The phases areseparated and the acidic solution reextracted with ethyl acetate. Theextracts of the acidified aqueous solution are combined, dried, andevaporated to give the N-t-butoxycarbonyl derivative of7-(α-amino-p-ureidophenylacetamido)cephalosporanic acid.

A mixture of the above product (2.77 g, 4.93 mmol) in pH 6.4 phosphatebuffer (30 ml) is treated with NaHCO₃ (1.085 g, 12.33 mmol) and then4-mercapto-1,2,3-triazole (0.748 g, 7.4 mmol). The solution is warmed to70° and stirred at 70° ± 3° for 2.75 hours. The solution is cooled,filtered, and acidified to pH 2.5 producing a residue. The solvents aredecanted and the residue is washed with water. The product is dissolvedin ethyl acetate, washed with water, dried, and evaporated to theN-protected product.

The protected product is stirred at 0° to 5° with a 9:1 trifluoroaceticacid: anisole solution for 70 minutes. The solvents are evaporated andthe residue is poured with rapid stirring into ether (350 ml). Thetrifluoroacetate salt of the product is collected, washed with ether,dissolved in water, and then stirred with basic ion-exchange resin. Theaqueous solution is lypholized to give the product.

EXAMPLE 6

When an equimolar amount of the following7-amino-3-heterocyclicthiomethyl-3-cephem-4-carboxylic acids aresubstituted for7-amino-3-(5-methyl-1,3,4-thiadiazol-2-ylthiomethyl)-3-cephem-4-carboxylicacid in the procedure of Example 2, the corresponding7-(α-amino-p-ureidophenylacetamido)-3-heterocyclicthiomethyl-3-cephem-4-carboxylic acid isformed.

7-Amino-3-(tetrazol-5-ylthiomethyl)-3-cephem-4-carboxylic acid

7-Amino-3-(1,3,4-thiadiazol-2-ylthiomethyl)-3-cephem-4-carboxylic acid

7-Amino-3-(5-trifluoromethyl-1,3,4-thiadiazol-2-ylthiomethyl)-3-cephem-4-carboxylicacid

7-Amino-3-(5-ethyl-1,3,4-thiadiazol-2-ylthiomethyl)-3-cephem-4-carboxylicacid

7-Amino-3-(5-n-butyl-1,3,4-thiadiazol-2-ylthiomethyl)-3-cephem-4-carboxylicacid

7-Amino-3-(5-dimethylamino-1,3,4-thiadiazol-2-ylthiomethyl)-3-cephem-4-carboxylicacid

7-Amino-3-(5-mercapto-1,3,4-thiadiazol-2-ylthiomethyl)-3-cephem-4-carboxylicacid

7-Amino-3-(3-methylthio-1,2,4-thiadiazol-5-ylthiomethyl)-3-cephem-4-carboxylicacid

7-Amino-3-(3-methyl-1,2,4-thiadiazol-5-ylthiomethyl)-3-cephem-4-carboxylicacid

7-Amino-3-(1,2,4-triazol-3-ylthiomethyl)-3-cephem-4-carboxylic acid

7-Amino-3-(5-methyl-1,2,4-triazol-3-ylthiomethyl)-3-cephem-4-carboxylicacid

7-Amino-3-(4-methyl-1,2,4-triazol-3-ylthiomethyl)-3-cephem-4-carboxylicacid

7-Amino-3-(4,5-dimethyl-1,2,4-triazol-3-ylthiomethyl)-3-cephem-4-carboxylicacid

7-Amino-3-(2,5-dimethyl-1,2,4-triazol-3-ylthiomethyl)-3-cephem-4-carboxylicacid

7-Amino-3-(4-methyl-5-trifluoromethyl-1,2,4-triazol-3-ylthiomethyl)-3-cephem-4-carboxylicacid

7-Amino-3-(5-ethyl-1,2,4-triazol-3-ylthiomethyl)-3-cephem-4-carboxylicacid

7-Amino-3-(4-ethyl-1,2,4-triazol-3-ylthiomethyl)-3-cephem-4-carboxylicacid

7-Amino-3-(1-methyl-1,2,4-triazol-5-ylthiomethyl)-3-cephem-4-carboxylicacid

7-Amino-3-(1-methyl-1,2,4-triazol-3-ylthiomethyl)-3-cephem-4-carboxylicacid

7-Amino-3-(1-ethyl-1,2,4-triazol-5-ylthiomethyl)-3-cephem-4-carboxylicacid

7-Amino-3-(1-ethyl-1,2,4-triazol-3-ylthiomethyl)-3-cephem-4-carboxylicacid

7-Amino-3-(4-allyl-1,2,4-triazol-3-ylthiomethyl)-3-cephem-4-carboxylicacid

7-Amino-3-(5-methoxymethyl-1,2,4-triazol-3-ylthiomethyl)-3-cephem-4-carboxylicacid

7-Amino-3-(5-cyclopropyl-1,2,4-triazol-3-ylthiomethyl)-3-cephem-4-carboxylicacid

7-Amino-3-(5-bromo-1,2,4-triazol-3-ylthiomethyl)-3-cephem-4-carboxylicacid

7-Amino-3-(5-hydroxy-4-methyl-1,2,4-triazol-3-ylthiomethyl)-3-cephem-4-carboxylicacid

7-Amino-3-(5-hydroxy-4-ethyl-1,2,4-triazol-3-ylthiomethyl)-3-cephem-4-carboxylicacid

7-Amino-3-(5-hydroxy-1,2,4-triazol-3-ylthiomethyl)-3-cephem-4-carboxylicacid

7-Amino-3-(1-methyl-1,2,3-triazol-5-ylthiomethyl)-3-cephem-4-carboxylicacid

7-Amino-3-(4-methyl-1,2,3-triazol-5-ylthiomethyl)-3-cephem-4-carboxylicacid

7-Amino-3-(1,3,4-oxadiazol-2-ylthiomethyl)-3-cephem-4-carboxylic acid

7-Amino-3-(5-methyl-1,3,4-oxadiazol-2-ylthiomethyl)-3-cephem-4-carboxylicacid

7-Amino-3-(4-pyrimidylthiomethyl)-3-cephem-4-carboxylic acid

7-Amino-3-(2-pyrazinylthiomethyl)-3-cephem-4-carboxylic acid

7-Amino-3-(3-pyridylthiomethyl)-3-cephem-4-carboxylic acid

7-Amino-3-(4-pyridylthiomethyl)-3-cephem-4-carboxylic acid

7-Amino-3-(1-oxide-2-pyridylthiomethyl)-3-cephem-4-carboxylic acid.

EXAMPLE 7 α-t-Butoxycarbonylamino-m-ureidophenylacetic acid

A mixture of m-nitrophenylglycine (58.0 g, 0.296 mol), MgO (23.92 g, 0.6mol) and t-butoxylcarbonyl azide (85.2 g, 0.6 mol) in 50% aqueousdioxane (700 ml) was heated at 45° for 17 hours. The reaction wascooled, water (750 ml) was added and the mixture was filtered. Thefiltrate was washed with ether (1000 ml), acidified to pH 3 with 40% H₃PO₄, and extracted with ethyl acetate. The dried extracts wereevaporated to give α-t-butoxycarbonylamino-m-nitrophenylacetic acid(70.3 g).

The above product (50.0 g, 0.17 mol) was dissolved in methanol (900 ml)and hydrogenated at 50 psi for 1 hour with 5% palladium on carbon (4 g).Evaporation of the filtered reaction solution gave the m-amino product(44.8 g).

A solution of α-t-butoxycarbonylamino-m-aminophenylacetic acid (2.66 g,0.01 mol) and potassium cyanate (0.81 g, 0.01 mol) in a mixture of water(35 ml) and acetic acid (2.5 ml) was heated at 56° for 3 hours. Thesolution was cooled, acidified with 40% H₃ PO₄, and extracted with ethylacetate. The dried extracts were evaporated to give the title product:mp 93°-95° (dec).

EXAMPLE 8 D-α-t-Butoxycarbonylamino-p-(3-methylureido)phenylacetic acid

Methyl isocyanate (0.314 g, 5.5 mmol) was added to a solution ofα-t-butoxycarbonylamino-p-aminophenylacetic acid (1.35 g, 5 mmol) inacetone (100 ml). The reaction was stirred at room temperature for onehour. More methyl isocyanate (0.2 g) was added and the solution wasstirred an additional two hours. The solvent was evaporated and theresidue was dissolved in water containing enough NaHCO₃ to maintain pH8. The basic solution was washed with ethyl acetate, acidified to pH 1.5with 3N HCl, and extracted with ethyl acetate. The dried extracts wereevaporated to give the title product.

EXAMPLE 9 D-α-t-Butoxycarbonylamino-p-(3,3-dimethylureido)phenylaceticacid

Dimethylcarbamoyl chloride (1.07 g, 0.01 mol) is added slowly to asolution of D-α-t-butoxycarbonylamino-p-aminophenylacetic acid (2.66 g,0.01 mol) in 50 ml of dioxane containing triethylamine (2.02 g, 0.02mol). After stirring at room temperature for several hours the mixtureis concentrated to half volume, diluted with water and the pH adjustedto 8. After extraction with ether the aqueous phase is adjusted to pH 2and extracted with ethyl acetate. Evaporation of the dried ethyl acetateextracts in vacuo gave the desired product.

EXAMPLE 10 α-t-Butoxycarbonylamino-p-thioureidophenylacetic acid

To a suspension of α-acetamido-p-aminophenylacetic acid (1.04 g, 5 mmol)in water (5 ml) was added 10% NaOH until solution was effected. Thesolution was diluted with an equal volume of ethanol and then t-butylisothiocyanate (1.64 g, 15 mmol) in ethanol was added. The reaction wasstirred at room temperature overnight and then refluxed 24 hours. Theethanol was evaporated in vacuo and the aqueous phase was acidified andfiltered. The aqueous filtrate was extracted with ethyl acetate. Thedried extracts were evaporated to giveα-acetamido-p-(3-t-butylthioureido)phenylacetic acid.

The above product (0.1 g, 0.31 mmol) was heated with 12N HCl (1 ml) on asteam bath for 15 minutes after gas evolution stopped. The solution wasdiluted with water and concentrated in vacuo. The residue was mixed withwater and concentrated an additional three times and then trituratedwith isopropanol to give the title product.

EXAMPLE 113-(5-Methyl-1,3,4-thiadiazol-2-ylthiomethyl)-7-(α-amino-m-ureidophenylacetamido)-3-cephem-4-carboxylicacid

To a solution of t-butyl7-amino-3-(5-methyl-1,3,4-thiadiazol-2-ylthiomethyl)-3-cephem-4-carboxylate(1.2 g, 3 mmol) and α-t-butoxycarbonylamino-m-ureidophenylacetic acid(0.93 g, 3 mmol) in tetrahydrofuran (75 ml) was addeddicyclohexylcarbodiimide (0.61 g, 3 mol). The mixture was stirred atroom temperature for 3 hours and then the solid urea was filtered off.The filtrate was evaporated in vacuo and the residue was triturated withether. The solid product was chromatographed on silica gel using 50:50benzene: acetone as eluent.

The t-butyl ester (0.88 g) was stirred with trifluoroacetic acid (12 ml)for 1 hour at room temperature. The solution was evaporated in vacuo ata temperature below 30°. The residue was triturated with ether to givethe trifluoroacetate salt of the title compound: 0.75 g, mp 158-165(dec). The zwitterionic product is obtained from the salt by standardmethods such as described in Example 2.

EXAMPLE 123-(1,2,3-Triazol-4-ylthiomethyl)-7-(α-amino-m-ureidophenylacetamido)-3-cepham-4-carboxylicacid

When 7-ACA, 4-mercapto-1,2,3-triazole, andα-t-butoxycarbonylamino-m-ureidophenylacetic acid are reacted accordingto the procedure of Example 5 the title compound is obtained.

EXAMPLE 133-(1-Methyltetrazol-5-ylthiomethyl)-7-(α-amino-m-ureidophenylacetamido)-3-cephem-4-carboxylicacid

Using α-t-butoxycarbonylamino-m-ureidophenylacetic acid in the procedureof Example 4, the title compound was obtained: mp 158°-165°(trifluoroacetic acid salt).

EXAMPLE 143-Methyl-7-(α-amino-m-ureidophenylacetamido)-3-cephem-4-carboxylic acid

The title compound was prepared according to the procedure of Example 3using α-t-butoxycarbonylamino-m-ureidophenylacetic acid and t-butyl7-aminodesacetoxycephalosporanic acid.

EXAMPLE 15

When the 7-aminocephem compounds enumerated in Example 6 are acylatedwith α-t-butoxycarbonylamino-m-ureidophenylacetic acid and the aminogroup is subsequently deblocked according to the procedure of Example 2,the corresponding 7-(α-amino-m-ureidophenylacetamido)cephalosporincompound is obtained.

EXAMPLE 163-(1-Methyltetrazol-5-ylthiomethyl)-7-[α-amino-p-(3-methylureido)phenylacetamido]-3-cephem-4-carboxylicacid

To a solution of α-t-butoxycarbonylamino-p-(3-methylureido)phenylaceticacid (2.1 g, 6.5 mmol) in dry tetrahydrofuran (75 ml) was addedtriethylamine (0.89 ml) and N-methylmorpholine (3 drops) causing a solidto precipitate. The suspension was cooled to -10° and isobutylchloroformate (0.84 ml) was added. The reaction was stirred at -10° for45 minutes and then a solution of7-amino-3-(1-methyltetrazol-5-ylthiomethyl)-3-cephem-4-carboxylic acid(2.13 g, 6.5 mmol) in a mixture of 50% aqueous tetrahydrofuran (25 ml)and triethylamine (0.89 ml) was added over a 15 minute period. Thereaction was stirred at ice bath temperature for one hour and then atroom temperature one hour. The organic solvents were evaporated and theresidue was dissolved in water (150 ml). The aqueous phase was washedwith ethyl acetate, adjusted to pH 2, and extracted with ethyl acetate(800 ml). The dried extracts were concentrated to a volume of 100 ml andthe N-protected product was collected, 1.63 g.

The above product (1.2 g) was added to ice cold trifluoracetic acid (10ml) and stirred after cooling for 8 minutes. The solution was partiallyevaporated and then diluted with ether. The trifluoroacetate salt wascollected. The salt was dissolved in water (100 ml) and treated withbasic ion-exchange resin (Amberlite IR-45) until pH 5.4 was obtained.After filtration the aqueous solution was lypholized to give the titleproduct (0.44 g).

EXAMPLE 17

Acylation of 7-ADCA,7-amino-3-(5-methyl-1,3,4-thiadiazol-2-ylthiomethyl)-3-cephem-4-carboxylicacid or any of the cephem compounds named in Example 6 withα-t-butoxycarboxyamino-p-(3-methylureido)phenylacetic acid followed bydeblocking according to the procedure of Example 2 gives thecorresponding 7-[α-amino-p-(3-methylureido)-phenylacetamido]cephemcompound. 7-[α-amino-p-(3-methylureido)phenylacetamido]-3-(1,2,3-triazol-4-ylthiomethyl)-3-cephem-4-carboxylicacid is prepared by reacting 7-ACA withα-t-butoxycarbonylamino-p-(3-methylureido)phenylacetic acid and thenwith 4-mercapto-1,2,3-triazole with subsequent deblocking of the aminogrouping according to the procedure of Example 5.

EXAMPLE 18

Using α-t-butoxycarbonylamino-p-(3,3-dimethylureido) phenylacetic acidto acylate the 7-aminocephem compounds of Examples 2,3,4 and 6 accordingto the procedure of Example 2 gives the appropriate7-[α-amino-p-(3,3-dimethylureido)phenylacetamido]cephalosporanic acids.Using N-t-butoxycarbony-p-(3,3-dimethylureido)phenylglycine as theacylating agent in the procedure of Example 5 gives7-[α-amino-p-(3,3-dimethylureido)phenylacetamido]-3-(1,2,3-triazol-4-ylthiomethyl)-3-cephem-4-carboxylicacid as the product.

EXAMPLE 19

When7-amino-3-(5-methyl-1,3,4-thiadiazol-2-ylthiomethyl)-3-cephem-4-carboxylicacid is reacted with α-t-butoxycarbonylamino-p-thioureidophenylaceticacid according to the procedure of Example 2,3-(5-methyl-1,3,4-thiadiazol-2-ylthiomethyl)-7-(α-amino-p-thioureidophenylacetamido)-3-cephem-4-carboxylicacid is obtained.

EXAMPLE 20

When 7-ACA, α-t-butoxycarbonylamino-p-thioureidophenylacetic acid, and4-mercapto-1,2,3-triazole are reacted by the procedure of Example 5,3-(1,2,3-triazol-4-ylthiomethyl)-7-(α-amino-p-thioureidophenylacetamido)-3-cephem-4-carboxylicacid is obtained.

EXAMPLE 21

When α-t-butoxycarbonylamino-p-thioureidophenylacetic acid is used toacylate any of the 7-aminocephem compounds listed in Examples 3, 4, and6, the corresponding7-(α-amino-p-thioureidophenylacetamido)cephalosporanic acids areobtained.

EXAMPLE 22 α-t-Butoxycarbonylamino-p-methylaminophenylacetic acid

Method A.

A mixture of α-t-butoxycarbonylamino-p-aminophenylacetic acid (26.6 g,0.1 mol), formaldehyde (8.1 ml of a 37% aq. solution) and phthalimide(14.7 g, 0.1 mol) in 200 ml of absolute ethanol is heated at reflux for1 hour. After cooling to room temperature Raney nickel catalyst is addedand the mixture is hydrogenated under 60 psi of H₂ at 100° for 2 hours.The catalyst is filtered off and the solvent is evaporated in vacuo.Following chromatography on silica gel the desired product is obtained.

Method B.

Dimethylsulfate (1.26 g, 0.01 mol) is added gradually to a solution ofα-t-butoxycarbonylamino-p-aminophenylacetic acid (13.3 g, 0.05 mol) in50 ml of 50% aqueous methanol containing NaOH (2.4 g, 0.06 mol). Whenthe reaction was complete the reaction mixture was acidified andextracted with ethyl acetate. The desired product was isolated by columnchromatography on silica gel.

EXAMPLE 23 α-t-Butoxycarbonylamino-p-(1-methylureido)phenylacetic acid

α-t-Butoxycarbonylamino-p-methylaminophenylacetic acid is reacted withpotassium cyanate according to the procedure in Example 1 to give thetitle compound.

EXAMPLE 243-(1,2,3-Triazol-4-ylthiomethyl)-7-[α-amino-p-(1-methylureido)phenylacetamido]-3-cephem-4-carboxylicacid

When α-t-butoxycarbonylamino-p-(1-methylureido)phenylacetic acid isreacted with 7-ACA and 4-mercapto-1,2,3-triazole according to theprocedure of Example 5, the title compound is obtained.

EXAMPLE 25

Acylation of any of the 7-aminocephem compounds enumerated in Examples2, 3, 4 or 6 with α-t-butoxycarbonylamino-p-(1-methylureido)phenylaceticacid according to the procedure of Example 2 gives the appropriate7-[α-amino-p-(1-methylureido)phenylacetamido]-3-cephem compound.

EXAMPLE 26

When 7-ACA is acylated withα-t-butoxycarbonylamino-p-(3-methylureido)phenylacetic acid orα-t-butoxycarbonylamino-p-(3,3-dimethylureidophenylacetic acid and theproduct treated with 4-mercapto-1,2,3-triazole, all according to theprocedure of Example 5, the corresponding3-(1,2,3-triazol-4-ylthiomethyl)-7-(p-substitutedureidophenylacetamido)-3-cephem-4-carboxylicacid is obtained.

EXAMPLE 27

When the N-t-butoxycarbonyl derivative of7-(α-amino-p-ureidophenylacetamido)cephalosporanic acid from Example 5was deblocked according to the procedure in Example 2,7-(α-amino-p-ureidophenylacetamido)cephalosporanic acid was obtained.

When 7-ACA was acylated withα-t-butoxycarbonylamino-m-ureidophenylacetic acid according to theprocedure of Example 5 and then deblocked7-(α-amino-p-ureidophenylacetamido)cephalosporanic acid was obtained.

EXAMPLE 28

Following the procedure of Example 2, 7-ACA is reacted with theN-t-butoxycarbonyl derivatives of α-amino-p-(3-methylureido)phenylaceticacid, α-amino-p-(3,3-dimethylureido)phenylacetic acid,α-amino-p-(1-methylureido)phenylacetic acid, orα-amino-p-thioureidophenylacetic acid to give the corresponding7-(α-amino-p-substituted ureidophenylacetamido)cephalosporanic acid.

EXAMPLE 29

Acylation of 7-amino-3-methylthiomethyl-3-cephem-4-carboxylic acidaccording to the procedure of Example 2 with the N-protected derivativeof α-amino-p-ureidophenylacetic acid, α-amino-m-ureidophenylacetic acid,α-amino-p-(3-methylureido)phenylacetic acid,α-amino-p-(3,3-dimethylureido)phenylacetic acid,α-amino-p-(1-methylureido)phenylacetic acid, orα-amino-p-thioureidophenylacetic acid gives the corresponding7-(α-amino-α-ureidophenylacetamido)-3-methylthiomethyl-3-cephem-4-carboxylicacid.

EXAMPLE 30

When 7-amino-3-methoxymethyl-3-cephem-4-carboxylic acid is acylated bythe procedure of Example 2 with the acetic acid compounds listed inExample 29 the corresponding7-(α-amino-α-ureidophenylacetamido)-3-methoxymethyl-3-cephem-4-carboxylicacid is obtained.

EXAMPLE 31

An injectable pharmaceutical composition is prepared by dissolving 500mg of sodium3-(5-methyl-1,3,4-thiadiazol-2-ylthiomethyl)-7-(α-amino-p-ureidophenylacetamido)-3-cephem-4-carboxylatein sterile water or sterile normal saline solution (1-2 ml). Othercephalosporins of this invention are formulated in a similar manner.

An antibacterial capsule is comprised of the following components:

    ______________________________________                                        cephalosporin         500    mg                                               lactose               250    mg                                               magnesium stearate    75     mg                                               ______________________________________                                    

What is claimed is:
 1. A compound of the formula ##STR3## where: the R₂NCXNR group is at the para or meta position;R is hydrogen or lower alkylof 1-4 carbon atoms; X oxygen or sulfur; and Y is a easily removableprotecting group.
 2. A compound as claimed in claim 1 where Y ist-butoxycarbonyl, trichloroethoxycarbonyl, benzyloxycarbonyl, or themethyl acetoacetate adduct.
 3. A compound as claimed in claim 2 where Ris hydrogen or methyl.
 4. A compound as claimed in claim 3 being thecompound α-t-butoxycarbonylamino-p-ureidophenylacetic acid.
 5. Acompound as claimed in claim 3 being the compoundα-t-butoxycarbonylamino-m-ureidophenylacetic acid.
 6. A compound asclaimed in claim 3 being the compoundα-t-butoxycarbonylamino-p-thioureidophenylacetic acid.
 7. A compound asclaimed in claim 3 being the compoundα-t-butoxycarbonylamino-m-thioureidophenylacetic acid.
 8. A compound asclaimed in claim 3 being the compoundα-t-butoxycarbonylamino-p-(3-methylureido)phenylacetic acid.
 9. Acompound as claimed in claim 3 being the compoundα-t-butoxycarbonylamino-p-(3,3-dimethylureido)-phenylacetic acid.
 10. Acompound as claimed in claim 3 being the compoundα-t-butoxycarbonylamino-p-(1-methylureido)phenylacetic acid.